The outer blood-retinal barrier is formed in part by the tight junctions of the retinal pigment epithelium (RPE). This blood- tissue barrier separates the photosensitive retina from the underlying vascular choriocapillaris. Maintenance of these tight junctions is vital to normal retinal physiology as observed in some retinopathies that result from a breakdown of this barrier. Tight junctions are composed of both transmembrane proteins and a submembranous complex of proteins known as the zonula occludens, or ZO, proteins. The ZO proteins belong to a larger class of proteins, the MAGUKs, that contain globular domains termed PDZ domain. Little is known about the function of the PDZ domain, although it has been suggested that they mediate interactions between the ZOs and may play a role in tight junction regulation. This proposal examines the role of ZO-2 subdomains in the regulation of RPE tight junctions using recombinant DNA techniques. A unique culture model (Ban and Rizzolo, 1997) that allows for different aspects of tight junction function to be assayed will be used to study the effect of gain-of-function and loss-of-function mutants in vitro.